Neoplasia, or a process of rapid cellular proliferation resulting in new, abnormal growth, is a characteristic of many diseases which can be serious, and sometimes, life-threatening. Typically, neoplastic growth of cells and tissues is characterized by greater than normal proliferation of cells, wherein the cells continue to grow even after the instigating factor (e.g., tumor promoter, carcinogen, virus) is no longer present. The cellular growth tends to show a lack of structural organization and/or coordination with the normal tissue and usually creates a mass of tissue (e.g., a tumor) which may be benign or malignant. Malignant cellular growth, or malignant tumors (cancer), are a leading cause of death worldwide, and the development of effective therapy for neoplastic disease is the subject of a large body of research. Although a variety of innovative approaches to treat and prevent cancers have been proposed, many cancers continue to cause a high rate of mortality and may be difficult to treat or relatively unresponsive to conventional therapies. In addition, patients may respond differently to various cancer therapies, making some approaches useful for some patients and not for others. Therefore, there is a continuing need in the art for the identification of additional cancer risk factors and methods for early diagnosis and therapy for cancers, as well as methods for identifying patients that are expected to benefit from a particular type of therapy.
Illustrating this point, non-small cell lung cancer (NSCLC) is the leading cause of cancer death in the world. While chemotherapy has produced modest survival benefits in advanced stages, standard two-drug combinations generate considerable toxicity and require intravenous administration (Non-small Cell Lung Cancer Collaborative Group, 1995; Schiller et al., 2002; Kelly et al., 2001). Progress in the field of lung cancer biology led to the development of small molecule inhibitors of target proteins involved in the proliferation, apoptosis and angiogenesis. Targeted therapy agents such as imatinib and trastuzumab produced consistent survival benefit in chronic myeloid leukemia (Druker, 2001), gastrointestinal stromal tumors (GIST) (Demetri 2002) and breast cancers that overexpress the target proteins (Slamon 2001). The epidermal growth factor receptor (EGFR) superfamily, including the four distinct receptors EGFR/erbB-1, HER2/erbB-2, HER3/erbB-3, and HER4/erbB-4, was early identified as a potential therapeutic target in solid tumors. After ligand binding, these receptors homo- and heterodimerize, and the tyrosine-kinase domain is activated, initiating a cascade of events implicated in the development and progression of cancer through effects on cell-cycle progression, apoptosis, angiogenesis, and metastasis (Salomon et al., 2001; Arteaga, 2002; Hirsch et al., 2003, Lung Cancer; Ciardello and Tortora, 2001). EGFR is overexpressed in many human epithelial malignancies, including NSCLC (Hirsch et al., 2003, J. Clin. Oncol.; Salomon et al., 1995).
Given the biological importance of the EGFR molecular network in carcinomas, several molecules were synthesized to inhibit the tyrosine kinase domain of EGFR (Levitzki and Gazit, 1995; Levitt and Koty, 1999). Among the most promising of these new drugs are gefitinib (ZD 1839, Iressa®, AstraZeneca, UK), and erlotinib (OSI 774, Tarceva®, Genentech, USA). Both are orally active, selective EGFR tyrosine-kinase inhibitors (EGFR-TKI) that demonstrated antitumor activity against a variety of human cancer cell lines expressing EGFR (Ciardiello et al., 2000). Likewise, both have well documented activity as single agents in phase I studies, including chemotherapy resistant NSCLC patients who had response rates of about 10% (Kris et al., 2000, Lung Cancer; Baselga et al., 2002; Herbst et al, 2002; Ranson et al., 2002; Hidalgo et al., 2001). Activity was confirmed in large phase II trials showing response rates of 19-26% in previously untreated, advanced NSCLC patients, and 12-18% in patients who had failed one or more prior chemotherapy combinations (Fukuoka et al., 2003; Kris et al., 2003, JAMA; Perez-Soler et al., 2001; Miller et al., 2003). More recently, a phase III trial (BR21) comparing erlotinib with placebo as a second or third line therapy reported a survival benefit for the EGFR inhibitor (Hazard Ratio: 0.73) (Shepherd et al., 2004). Importantly, this survival benefit was not confined to objective responders, nor to a single gender or histology, which makes selection based on clinical and histopathological features alone difficult.
In phase II trials with gefitinib, no correlation was detected between EGFR protein expression and response to therapy, although few studies have directly addressed this question. Patients with squamous cell carcinomas had lower response rates compared to patients with adenocarcinoma despite their higher rates of EGFR expression (Ciardiello et al., 2000; Fukuoka et al., 2003; Kris et al., 2003, JAMA). Recent reports showed that specific missense and deletion mutations in the tyrosine kinase domain of the EGFR gene (Lynch et all, 2004; Paez et al., 2004) are significantly associated with gefitinib sensitivity. However, while objective response has been reported in up to 18% and symptomatic improvement in 40% of the unselected gefitinib treated NSCLC patients (Fukuoka et al., 2003; Kris et al., 2003, JAMA), the frequency of these mutations in unselected US patients is low (Paez et al., 2004). These observations and the finding that objective response can be detected in patients carrying apparently wild type allele of the EGFR gene (Lynch et al, Pao et al., Han et al (JCO, 23:2493, 2005), Mitsudomi et al., JCO 23:2513, 2005, Kim et al., Clinical Cancer Res, 11:2244, 2005) suggest that other mechanisms are also involved in the response to gefitinib. Furthermore, while these activating mutations identify patients with high response rates, they cannot account for the high stable disease rates, reported to occur in about 30% of NSCLC patients treated with gefitinib (Fukuoka et al., 2003; Kris et al., 2003, JAMA).
In summary, there are no reliable selection criteria for determining which cancer patients, including NSCLC patients, will benefit from treatment with EGFR inhibitors exemplified by, but not limited to, gefitinib. Therefore, it is of great interest to identify both patients that would benefit from EGFR inhibitors and patients who are not going to benefit from such therapy, as well as to identify treatments which can improve the responsiveness of cancer cells which are resistant to EGFR inhibitors, and to develop adjuvant treatments that enhance the response.